Matt:LabNotes/2015-7-21

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Design 3 New RNA FISH Probes for Validating DARTFISH in Motor Neurons[edit]

  • Current RNA FISH probes don't include a high expression gene in Sebastian's motor neurons

Pick 3 new genes that validate DARTFISH[edit]

  • Validate normalization method can properly characterize true gene expression
  • Pick only genes with >10 DARTFISH raw count in 8 positions (see above data link)
  1. High DARTFISH raw count and high DARTFISH normalized value
    • KIF5C: #3 in raw count and #7 in gDNA normalized value
  2. High DARTFISH raw count and low DARTFISH normalized value
    • PHKB: #15 in raw count and #77 in gDNA normalized value
  3. Low DARTFISH raw count and high DARTFISH normalized value
    • OLFM1: #78 in raw count and #12 in gDNA normalized value

mRNA coding sequence[edit]

  • Find using UCSC Genome Browser 'UCSC Genes' track to pick shortest transcript variant that shares as many exons with other variants

KIF5C[edit]

  • Picked short transcript at 3' end

Description: Homo sapiens kinesin family member 5C (KIF5C), mRNA. mRNA sequence from here Transcript (Including UTRs)

  Position: chr2:149,856,449-149,883,273 Size: 26,825 Total Exon Count: 7 Strand: +

Coding Region

  Position: chr2:149,856,881-149,868,190 Size: 11,310 Coding Exon Count: 6  

PHKB[edit]

  • Picked short transcript at 3' end

Description: Homo sapiens phosphorylase kinase, beta (PHKB), transcript variant 2, mRNA. mRNA sequence from here Transcript (Including UTRs)

  Position: chr16:47,699,183-47,735,434 Size: 36,252 Total Exon Count: 7 Strand: +

Coding Region

  Position: chr16:47,699,794-47,733,277 Size: 33,484 Coding Exon Count: 7

OLFM1[edit]

  • Picked 2nd to shortest transcript that has 3' end

Description: Homo sapiens olfactomedin 1 (OLFM1), transcript variant 1, mRNA. mRNA sequence from here Transcript (Including UTRs)

  Position: chr9:137,979,817-138,013,030 Size: 33,214 Total Exon Count: 6 Strand: +

Coding Region

  Position: chr9:137,979,836-138,012,024 Size: 32,189 Coding Exon Count: 6  

Stellaris® Probe Designer version 4.1[edit]

  • Account: mzcai (pswd same as labnotes)
  • Sequences saved in shopping cart
  • Design parameters:
    • Organism: human
    • Masking level: 5
    • Max number of probes: 48
    • Probe length: 20
    • Min spacing: 2